Plant Cell Rep (2008) 27:411–424
DOI 10.1007/s00299-007-0474-9
REVIEW
Transgenic approaches for abiotic stress tolerance in plants:
retrospect and prospects
Pooja Bhatnagar-Mathur Æ V. Vadez Æ
Kiran K. Sharma
Received: 20 August 2007 / Revised: 21 October 2007 / Accepted: 22 October 2007 / Published online: 20 November 2007
Ó Springer-Verlag 2007
Abstract Abiotic stresses including drought are serious
threats to the sustainability of crop yields accounting for
more crop productivity losses than any other factor in
rainfed agriculture. Success in breeding for better adapted
varieties to abiotic stresses depend upon the concerted
efforts by various research domains including plant and
cell physiology, molecular biology, genetics, and breeding.
Use of modern molecular biology tools for elucidating the
control mechanisms of abiotic stress tolerance, and for
engineering stress tolerant crops is based on the expression
of specific stress-related genes. Hence, genetic engineering
for developing stress tolerant plants, based on the introgression of genes that are known to be involved in stress
response and putative tolerance, might prove to be a faster
track towards improving crop varieties. Far beyond the
initial attempts to insert ‘‘single-action’’ genes, engineering
of the regulatory machinery involving transcription factors
has emerged as a new tool now for controlling the
expression of many stress-responsive genes. Nevertheless,
the task of generating transgenic cultivars is not only
limited to the success in the transformation process, but
also proper incorporation of the stress tolerance. Evaluation
of the transgenic plants under stress conditions, and
understanding the physiological effect of the inserted genes
at the whole plant level remain as major challenges to
overcome. This review focuses on the recent progress in
using transgenic technology for the improvement of abiotic
stress tolerance in plants. This includes discussion on the
Communicated by P. Kumar.
P. Bhatnagar-Mathur V. Vadez K. K. Sharma (&)
International Crops Research Institute for the Semi-Arid Tropics
(ICRISAT), Patancheru, Andhra Pradesh 502 324, India
e-mail: [email protected]
evaluation of abiotic stress response and the protocols for
testing the transgenic plants for their tolerance under closeto-field conditions.
Keywords Abiotic stress Drought tolerance Genetic engineering Transcription factors Transpiration efficiency
Introduction
Abiotic stresses adversely affect growth and productivity
and trigger a series of morphological, physiological, biochemical and molecular changes in plants. Drought,
temperature extremes, and saline soils are the most common abiotic stresses that plants encounter. Globally,
approximately 22% of the agricultural land is saline (FAO
2004), and areas under drought are already expanding and
this is expected to increase further (Burke et al. 2006).
Often crops are exposed to multiple stresses, and the
manner in which a plant senses and responds to different
environmental factors appears to be overlapping. Gene
expression profiles of either drought- or salt-stressed barley
plants indicated that although, various genes were differentially regulated in response to different stresses, they
possibly induce a similar defense response (Ozturk et al.
2002).
When a plant is subjected to abiotic stress, a number of
genes are turned on, resulting in increased levels of several
metabolites and proteins, some of which may be responsible for conferring a certain degree of protection to these
stresses. A key to progress towards breeding better crops
under stress has been to understand the changes in cellular,
biochemical and molecular machinery that occur in
response to stress. Modern molecular techniques involve
123
412
the identification and use of molecular markers that can
enhance breeding programs. However, the introgression of
genomic portions (QTLs) involved in stress tolerance often
brings along undesirable agronomic characteristics from
the donor parents. This is because of the lack of a precise
knowledge of the key genes underlying the QTLs. Therefore, the development of genetically engineered plants by
the introduction and/or overexpression of selected genes
seems to be a viable option to hasten the breeding of
‘‘improved’’ plants. Intuitively, genetic engineering would
be a faster way to insert beneficial genes than through
conventional or molecular breeding. Also, it would be the
only option when genes of interest originate from cross
barrier species, distant relatives, or from non-plant sources.
Indeed, there are several traits whose correlative association with resistance has been tested in transgenic plants.
Following these logical steps, various transgenic technologies have been used to improve stress tolerance in plants
(Allen 1995).
Stress-induced gene expression can be broadly categorized into three groups: (1) genes encoding proteins
with known enzymatic or structural functions, (2) proteins
with as yet unknown functions, and (3) regulatory proteins. Initial attempts to develop transgenics (mainly
tobacco) for abiotic stress tolerance involved ‘‘single
action genes’’ i.e., genes responsible for modification of a
single metabolite that would confer increased tolerance to
salt or drought stress Stress-induced proteins with known
functions such as water channel proteins, key enzymes for
osmolyte (proline, betaine, sugars such as trehalose, and
polyamines) biosynthesis, detoxification enzymes, and
transport proteins were the initial targets of plant transformation. In fact, metabolic traits, especially pathways
with relatively few enzymes, have been characterized
genetically and appear more amenable to manipulations
than structural and developmental traits. However, that
approach has overlooked the fact that abiotic stress tolerance is likely to involve many genes at a time, and that
single-gene tolerance is unlikely to be sustainable.
Therefore, a second ‘‘wave’’ of transformation attempts to
transform plants with the third category of stress-induced
genes, namely, regulatory proteins has emerged. Through
these proteins, many genes involved in stress response can
be simultaneously regulated by a single gene encoding
stress inducible transcription factor (Kasuga et al. 1999),
thus offering possibility of enhancing tolerance towards
multiple stresses including drought, salinity, and freezing.
It is interesting to note that this ‘‘second wave’’ has also
coincided with a better integration of genetic engineering
and plant physiology.
Further, genetic engineering allows controlling the
timing, tissue-specificity, and expression level of the
introduced genes for their optimal function. This is an
123
Plant Cell Rep (2008) 27:411–424
important consideration if the action of a given gene or
transcription factor is desired only at a specific time, in a
specific organ, or under specific conditions of stress. The
basic findings on stress promoters have led to a major shift
in the paradigm for genetically engineering stress-tolerant
crops in recent years (Katiyar et al. 1999). The most widely
used promoters in generating transgenic plants are constitutively expressed, i.e., they are turned on all the time and
throughout the plant life cycle. However, in cases where
the gene expression needs to be tailored to a specific organ
or a specific time, such constitutive promoters may not be a
suitable choice, especially for the stress-induced genes.
This is because the constitutive expression of some stressinduced genes may have serious deleterious effects on the
plant. Accordingly, the more recent efforts to generate
transgenic plants make use of gene cassettes driven by
stress-induced promoters. With an increasing number of
stress genes becoming available and genetic transformation
becoming more or less a routine procedure, characterization of stress-induced promoters (particularly those induced
by anaerobic, low or high temperature and salt stresses) has
taken a firm footing (Katiyar et al. 1999).
It is important to examine how transgenic plants are
evaluated, and how the proof-of-concept of gene effect in
model plants can be adapted to crop species. Unfortunately,
a substantial amount of published work involving the
assessment of transgenic plants under abiotic stresses has
shown effect of the transgene under growth environments
that are unlikely to occur in the natural conditions. So,
there is a need to set basic guidelines on the protocols to be
used to carry out a rigorous evaluation of the response of
transgenic plants to abiotic stresses. Since most of the work
carried out so far has focused on a few model plants, there
is also a need to document and summarize the major
achievements in crop plants.
This review summarizes the recent progress in using
transgenic plant technology for the improvement of abiotic
stress tolerance using examples from research targeted at
drought, salinity and temperature stresses, with particular
attention to how transgenic plants are evaluated.
Single action genes
Osmoprotectants
Severe osmotic stress causes detrimental changes in cellular
components. In stress-tolerant transgenic plants, many genes
involved in the synthesis of osmoprotectants—organic
compounds such as amino acids (e.g. proline), quaternary
and other amines (e.g. glycinebetaine and polyamines) and
a variety of sugars and sugar alcohols (e.g. mannitol,
trehalose and galactinol) that accumulate during osmotic
Plant Cell Rep (2008) 27:411–424
adjustment—have been used to date (Vincour and Altman
2005). Many crops lack the ability to synthesize the special
osmoprotectants that are naturally accumulated by stresstolerant organisms. It is believed that osmoregulation would
be the best strategy for abiotic stress tolerance, especially if
osmoregulatory genes could be triggered in response to
drought, salinity and high temperature. Therefore, a widely
adopted strategy has been to engineer certain osmolytes or
by over expressing such osmolytes in plants, as a potential
route to breed stress-tolerant crops.
Various strategies are being pursued to genetically
engineer osmoprotection in plants. The first step involved
in obtaining stress tolerant transgenic plants has been to
engineer genes that encode enzymes for the synthesis of
selected osmolytes (Bray 1993). This has resulted in a
profusion of reports involving osmoprotectants such as
glycine-betaine (Ishitani et al. 1997; Lilius et al. 1996;
Hayashi et al. 1997, 1998; Alia et al. 1998, 1999;
Sakamoto et al. 1998, 2000; Holmstrom 2000; McNeil
et al. 2000) and proline (Delauney and Verma 1993;
Nanjo et al. 1999a; Zhu et al. 1998; Yamada et al. 2005).
Also, a number of ‘‘sugar alcohols’’ (mannitol, trehalose,
myo-inositol and sorbitol) have been targeted for the
engineering of compatible-solute overproduction, thereby
protecting the membrane and protein complexes during
stress (Tarczynski et al. 1993; Yang et al. 1996; Shen
et al. 1997; Abebe et al. 2003; Holmstrom et al. 1996;
Zhao et al. 2000; Pilon-Smits et al. 1995, 1998, 1999;
Garg et al. 2002; Cortina and Culiáñez 2005; Gao et al.
2000). Similarly, transgenics engineered for the overexpression of polyamines have also been developed (Roy
and Wu 2001; 2002; Kumria and Rajam 2002; Waie and
Rajam 2003; Anderson et al. 1998; Capell et al. 2004).
Studies on the identification/isolation/cloning of genes
that are associated with improved flooding stress tolerance
have also focused on enzymes of the glycolytic and
alcohol fermentation pathways indicating that respiratory
pathway is affected in a major way in response to
anaerobic stress. Research on genetically altering the
levels of pdc and adh in tobacco and rice has been
extensively carried out to elucidate their role in submergence tolerance. Transgenic rice over- and underexpressing pyruvate decarboxylase 1 (pdc1) gene has also
been developed, which showed a positive correlation of
higher PDC activities with survival after submergence
(Quimlo et al. 2000).
The results of transgenic modifications of biosynthetic
and metabolic pathways in most of the above-mentioned
cases indicate that higher stress tolerance and the accumulation of compatible solutes may also protect plants
against damage by scavenging of reactive oxygen species
(ROS), and by their chaperone-like activities in maintaining protein structures and functions (Hare et al. 1998;
413
Bohnert and Shen 1999; McNeil et al. 1999; Diamant
et al. 2001). However, pleiotropic effects (e.g. necrosis
and growth retardation) have been observed due to disturbance in endogenous pathways of primary metabolisms.
Also, there are also some reports showing a negative effect
of osmotic stress on yield potential (Fukai and Cooper
1995). Genetic manipulations of compatible solutes do not
always lead to a significant accumulation of the compound
(except in some cases of proline over-production; Chen
and Murata 2002), thereby, suggesting that the function of
compatible solutes is not restricted to osmotic adjustment,
and that osmoprotection may not always confer drought
tolerance. A recent review (Serraj and Sinclair 2002)
shows that virtually none of the studies that tested the
effect of osmotic adjustment on yield under water stress
showed any benefit at all, since some benefit of osmotic
adjustment might be in the ability of plants to maintain
root growth under severe stress (Voetberg and Sharp
1991). Another recent study with chickpea has also shown
that osmotic adjustment provided no beneficial effect on
yield under drought stress (Turner et al. 2007). Besides,
the results of simulation modeling also suggest that
changes in a given metabolic process, (Passioura 1977,
2007), may end up with little benefit for actual yield under
stress (Sinclair et al. 2004). For agricultural practices,
over-synthesis of compatible solutes should not account
for the primary metabolic costs and hence to minimize the
pleiotropic effects, over-production of compatible solutes
should be stress-inducible and/or tissue specific (Garg
et al. 2002).
Detoxifying genes
In most of the aerobic organisms, there is a need to
effectively eliminate reactive oxygen species (ROS) generated as a result of environmental stresses. Depending on
the nature of the ROS, some are highly toxic and need to
be rapidly detoxified. In order to control the level of ROS
and protect the cells from oxidative injury, plants have
developed a complex antioxidant defense system to
scavenge the ROS. These antioxidant systems include
various enzymes and non-enzymatic metabolites that may
also play a significant role in ROS signaling in plants
(Vranova et al. 2002). A number of transgenic improvements for abiotic stress tolerance have been achieved
through detoxification strategy. These include transgenic
plants over expressing enzymes involved in oxidative
protection, such as glutathione peroxidase, superoxide
dismutase, ascorbate peroxidases and glutathione reductases (Zhu et al. 1999; Roxas et al. 1997). Transgenic
tobacco over expressing SOD in the chloroplast, mitochondria and cytosol have been generated (Bowler et al.
123
414
1991; Van Camp et al. 1996) and these have been shown
to enhance tolerance to oxidative stress induced by methyl
viologen (MV) in leaf disc assays. Overexpression of
chloroplast Cu/Zn SOD showed a dramatic improvement
in the photosynthetic performance under chilling stress
conditions in transgenic tobacco (Sen Gupta et al. 1993)
and potato plants (Perl et al. 1993). Tobacco transgenic
plants overexpressing MnSOD rendered enhanced tolerance to oxidative stress only in the presence of other
antioxidant enzymes and substrates (Slooten et al. 1995),
thereby, showing that the genotype and the isozyme
composition also have a profound effect on the relative
tolerance of the transgenic plants to abiotic stress (Rubio
et al. 2002). While transgenic alfalfa (Medicago sativa)
plants cv. RA3 overexpressing MnSOD in chloroplasts
showed lower membrane injury (McKersie et al. 1996),
the tobacco transgenic plants overproducing alfalfa aldose
reductase gene (MsALR) showed lower concentrations of
reactive aldehydes and increased tolerance against oxidative agents and drought stress (Oberschall et al. 2000).
Late embryogenesis abundant (LEA) proteins
LEA proteins represent another category of high molecular
weight proteins that are abundant during late embryogenesis and accumulate during seed desiccation and in
response to water stress (Galau et al. 1987). Amongst the
several groups of LEA proteins, those belonging to group 3
are predicted to play a role in sequestering ions that are
concentrated during cellular dehydration. These proteins
have 11-mer amino acid motifs with the consensus
sequence TAQAAKEKAGE repeated as many as 13 times
(Dure 1993). The group 1 LEA proteins are predicted to
have enhanced water-binding capacity, while the group 5
LEA proteins are thought to sequester ions during water
loss. Constitutive overexpression of the HVA1, a group 3
LEA protein from barley conferred tolerance to soil water
deficit and salt stress in transgenic rice plants (Xu et al.
1996). Constitutive or stress induced expression of the
HVA1 gene resulted in the improvement of growth characteristics and stress tolerance in terms of cell integrity in
wheat and rice under salt- and water-stress conditions
(Sivamani et al. 2000; Rohilla et al. 2002). Although, the
reported water use efficiency (WUE) was extremely low
when compared to other data reported in wheat cultigens,
transgenic rice (TNG67) plants expressing a wheat LEA
group 2 protein (PMA80) gene or the wheat LEA group 1
protein (PMA1959) gene resulted in increased tolerance to
dehydration and salt stresses (Cheng et al. 2002). Besides,
protective chaperone like function of LEA proteins acting
against cellular damage has been proposed (Vincour and
Altman 2005), indicating the role of LEA proteins in anti-
123
Plant Cell Rep (2008) 27:411–424
aggregation of enzymes under desiccation and freezing
stresses (Goyal et al. 2005).
Transporter genes
An important strategy for achieving greater tolerance to
abiotic stress is to help plants to re-establish homeostasis
under stressful environments, restoring both ionic and
osmotic homeostasis. This has been and continues to be a
major approach to improve salt tolerance in plants through
genetic engineering, where the target is to achieve Na+
excretion out of the root, or their storage in the vacuole. A
number of abiotic stress tolerant transgenic plants have
been produced by increasing the cellular levels of proteins
(such as vacuolar antiporter proteins) that control the
transport functions. For example, transgenic melon (Bordás
et al. 1997) and tomato (Gisbert et al. 2000) plants
expressing the HAL1 gene showed a certain level of salt
tolerance as a result of retaining more K+ than the control
plants under salinity stress.
A vacuolar chloride channel, AtCLCd gene, which is
involved in cation detoxification, and AtNHXI gene which is
homologous to NhxI gene of yeast have been cloned and
overexpressed in Arabidopsis to confer salt tolerance by
compartmentalizing Na+ ions in the vacuoles. Transgenic
Arabidopsis and tomato plants that overexpress AtNHX1
accumulated abundant quantities of the transporter in the
tonoplast and exhibited substantially enhanced salt tolerance
(Apse et al. 1999; Quintero et al. 2000; Zhang and Blumwald
2001). Salt Overly Sensitive 1 (SOS1) locus in A. thaliana,
which is similar to plasma membrane Na+/H+ antiporter
from bacteria and fungi, was cloned and overexpressed
using CaMV 35S promoter. The up-regulation of SOSI gene
was found to be consistent with its role in Na+ tolerance,
providing a greater proton motive force that is necessary for
elevated Na+/H+ antiporter activities (Shi et al. 2000).
Multifunctional genes for lipid biosynthesis
Transgenic approaches also aim to improve photosynthesis
under abiotic stress conditions through changes in the lipid
biochemistry of the membranes (Grover and Minhas 2000).
Adaptation of living cells to chilling temperatures is a function of alteration in the membrane lipid composition by
increased fatty acid unsaturation. Genetically engineered
tobacco plants over-expressing chloroplast glycerol-3-phosphate acyltransferase (GPAT) gene (involved in phosphatidyl
glycerol fatty acid desaturation) from squash (Cucurbita
maxima) and A. thaliana (Murata et al. 1992) showed an
increase in the number of unsaturated fatty acids and a corresponding decrease in the chilling sensitivity. Besides,
Plant Cell Rep (2008) 27:411–424
transgenic tobacco plants with silenced expression of chloroplast x3-fatty acid desaturase (Fad7, which synthesizes
trienoic fatty acids) were able to acclimate to high temperature as compared to the wild type (Murakami et al. 2000).
Heat shock protein genes
The heat shock response, the increased transcription of a set
of genes in response to heat or other toxic agent exposure is a
highly conserved biological response, occurring in all
organisms (Waters et al. 1996). The response is mediated by
heat shock transcription factor (HSF) which is present in a
monomeric, non-DNA binding form in unstressed cells and
is activated by stress to a trimeric form which can bind to
promoters of heat shock genes. The induction of genes
encoding heat shock proteins (Hsps) is one of the most
prominent responses observed at the molecular level of
organisms exposed to high temperature (Kimpel and Key
1985; Lindquist 1986; Vierling 1991).
Genetic engineering for increased thermo-tolerance by
enhancing heat shock protein synthesis in plants has been
achieved in a number of plant species (Malik et al. 1999; Li
et al. 2003; Katiyar-Agarwal et al. 2003). There have been
a few reports on positive correlations between the levels of
heat shock proteins and stress tolerance (Sun et al. 2001;
Wang et al. 2005). Although the precise mechanism by
which these heat shock proteins confer stress tolerance is
not known, a recent study demonstrated that in vivo
function of thermoprotection of small heat shock proteins
is achieved via their assembly into functional stress granules (HSGs; Miroshnichenko et al. 2005).
Regulatory genes
Many genes that respond to multiple stresses like dehydration and low temperature at the transcriptional level are
also induced by ABA (Mundy and Chua 1988), which
protects the cell from dehydration (Dure et al. 1989;
Skriver and Mundy 1990). In order to restore the cellular
function and make plants more tolerant to stress, transferring a single gene encoding a single specific stress protein
may not be sufficient to reach the required tolerance levels
(Bohnert et al. 1995). To overcome such constraints,
enhancing tolerance towards multiple stresses by a gene
encoding a stress inducible transcription factor that regulates a number of other genes is a promising approach
(Yamaguchi-Shinozaki et al. 1994; Chinnusamy et al.
2005). Therefore, a second category of genes of recent
preference for crop genetic engineering are those that
switch on transcription factors regulating the expression of
several genes related to abiotic stresses.
415
Transcription factors
An attractive target category for manipulation and gene
regulation is the small group of transcription factors that
have been identified to bind to promoter regulatory elements
in genes that are regulated by abiotic stresses (Shinozaki and
Yamaguchi-Shinozaki 1997; Winicov and Bastola 1997).
The transcription factors activate cascades of genes that act
together in enhancing tolerance towards multiple stresses.
Dozens of transcription factors are involved in the plant
response to drought stress (Vincour and Altman 2005;
Bartels and Sunkar 2005). Most of these fall into several
large transcription factor families, such as AP2/ERF, bZIP,
NAC, MYB, MYC, Cys2His2 zinc-finger and WRKY.
Individual members of the same family often respond differently to various stress stimuli. On the other hand, some
stress responsive genes may share the same transcription
factors, as indicated by the significant overlap of the geneexpression profiles that are induced in response to different
stresses (Seki et al. 2001; Chen and Murata 2002). Transcriptional activation of stress-induced genes has been
possible in transgenic plants over expressing one or more
transcription factors that recognize promoter regulatory
elements of these genes. Two families, bZIP and MYB, are
involved in ABA signaling and its gene activation. Many
ABA inducible genes share the (C/T) ACGTGGC consensus, cis-acting ABA-responsive element (ABRE) in their
promoter regions (Guiltinan et al. 1990; Mundy et al. 1990).
Introduction of transcription factors in the ABA signaling
pathway can also be a mechanism of genetic improvement of
plant stress tolerance. Constitutive expression of ABF3 or
ABF4 demonstrated enhanced drought tolerance in Arabidopsis, with altered expression of ABA/stress-responsive
genes, e.g. rd29B, rab18, ABI1 and ABI2 (Kagaya et al.
2002). Several ABA-associated phenotypes, such as ABA
hypersensitivity and sugar hypersensitivity, were observed
in such plants. Moreover, salt hypersensitivity was observed
in ABF3- and ABF4-overexpressing plants at the germination and young seedling stages indicating the possible
participation of ABF3 and ABF4 in response to salinity at
these particular developmental stages. Improved osmoticstress tolerance in 35S:At-MYC2/AtMYB2 transgenic
plants as judged by an electrolyte-leakage test was reported
by (Abebe et al. 2003). Transgenic Arabidopsis plants constitutively over-expressing a cold inducible transcription
factor (CBF1; CRT/DRE binding protein) showed tolerance
to freezing without any negative effect on the development
and growth characteristics (Jaglo-Ottosen et al. 1998). Overexpression of Arabidopsis CBF1 (CRT/DRE binding protein) has been shown to activate cor homologous genes at
non-acclimating temperatures (Jaglo et al. 2001). The CBF1
cDNA when introduced into tomato (Lycopersicon esculentum) under the control of a CaMV 35S promoter improved
123
416
tolerance to chilling, drought and salt stress but exhibited
dwarf phenotype and reduction in fruit set and seed number
(Hsieh et al. 2002). Another transcriptional regulator, Alfin1,
when overexpressed in transgenic alfalfa (Medicago sativa
L.) plants regulated endogenous MsPRP2 (NaCl-inducible
gene) mRNA levels, resulting in salinity tolerance, comparable, to a few available salt tolerant plants (Winicov and
Bastola 1999). Lee et al. (1995) produced thermo-tolerant
Arabidopsis plants by de-repressing the activity of ATHSF1,
a heat shock transcription factor leading to the constitutive
expression of heat shock proteins at normal temperature.
Several stress induced cor genes such as rd29A, cor15A, kin1
and cor6.6 are triggered in response to cold treatment, ABA
and water deficit stress (Thomashow 1998). There have been
numerous efforts in enhancing tolerance towards multiple
stresses such as cold, drought and salt stress in crops other
than the model plants like Arabidopsis, tobacco and alfalfa.
An increased tolerance to freezing and drought in Arabidopsis was achieved by overexpressing CBF4, a close CBF/
DREB1 homolog whose expression is rapidly induced during drought stress and by ABA treatment, but not by cold
(Haake et al. 2002). Similarly, a cis-acting element, dehydration responsive element (DRE) identified in A. thaliana,
is also involved in ABA-independent gene expression under
drought, low temperature and high salt stress conditions in
many dehydration responsive genes like rd29A that are
responsible for dehydration and cold-induced gene expression (Yamaguchi-Shinozaki and Shinozaki 1993; Iwasaki
et al. 1997; Nordin et al. 1991). Several cDNAs encoding the
DRE binding proteins, DREB1A and DREB2A have been
isolated from A. thaliana and shown to specifically bind and
activate the transcription of genes containing DRE sequences (Liu et al. 1998). DREB1/CBFs are thought to function in
cold-responsive gene expression, whereas DREB2s are
involved in drought-responsive gene expression. The transcriptional activation of stress-induced genes has been
possible in transgenic plants over-expressing one or more
transcription factors that recognize regulatory elements
of these genes. In Arabidopsis, the transcription factor
DREB1A specifically interacts with the DRE and induces
expression of stress tolerance genes (Shinozaki and
Yamaguchi-Shinozaki 1997). DREB1A cDNA under the
control of CaMV 35S promoter in transgenic plants elicits
strong constitutive expression of the stress inducible genes
and brings about increased tolerance to freezing, salt and
drought stresses (Liu et al. 1998). Strong tolerance to
freezing stress was observed in transgenic Arabidopsis
plants that overexpress CBF1 (DREB1B) cDNA under the
control of the CaMV 35S promoter (Jaglo-Ottosen et al.
1998). Subsequently, the overexpression of DREB1A has
been shown to improve the drought- and low-temperature
stress tolerance in tobacco, wheat and groundnut (Kasuga
et al. 2004; Pellegrineschi et al. 2004; Behnam et al. 2006;
123
Plant Cell Rep (2008) 27:411–424
Bhatnagar-Mathur et al. 2004, 2006). The use of stressinducible rd29A promoter minimized the negative effects on
plant growth in these crop species. However, overexpression of DREB2 in transgenic plants did not improve stress
tolerance, suggesting involvement of post-translational
activation of DREB2 proteins (Liu et al. 1998). Recently, an
active form of DREB2 was shown to transactivate target
stress-inducible genes and improve drought tolerance in
transgenic Arabidopsis (Sakuma et al. 2006). The DREB2
protein is expressed under normal growth conditions and
activated by osmotic stress through post-translational modification in the early stages of the osmotic stress response.
Another ABA-independent, stress-responsive and senescence-activated gene expression involves ERD gene, the
promoter analysis of which further identified two different
novel cis acting elements involved with dehydration stress
induction and in dark-induced senescence (Simpson et al.
2003). Similarly, transgenic plants developed by expressing a
drought-responsive AP2-type TF, SHN1-3 or WXP1, induced
several wax-related genes resulting in enhanced cuticular wax
accumulation and increased drought tolerance (Aharoni et al.
2004; Zhang et al. 2005). Thus, clearly, the overexpression of
some drought-responsive transcription factors can lead to the
expression of downstream genes and the enhancement of
abiotic stress tolerance in plants (see review, Zhang et al.
2004). The regulatory genes/factors reported so far not only
play a significant role in drought and salinity stresses, but also
in submergence tolerance. More recently, an ethyleneresponse-factor-like gene Sub1A, one of the cluster of three
genes at the Sub1 locus have been identified in rice and the
overexpression of Sub1A-1 in a submergence-intolerant
variety conferred enhanced submergence tolerance to the
plants (Xu et al. 2006), thus confirming the role of this gene in
submergence tolerance in rice.
Signal transduction genes
Genes involved in stress signal sensing and a cascade of
stress-signaling in A. thaliana has been of recent research
interest (Winicov and Bastola 1997; Shinozaki and
Yamaguchi-Shinozaki 1999). Components of the same
signal transduction pathway may also be shared by various
stress factors such as drought, salt and cold (Shinozaki and
Yamaguchi-Shinozaki 1999). Although there are multiple
pathways of signal-transduction systems operating at the
cellular level for gene regulation, ABA is a known component acting in one of the signal transduction pathways,
while others act independently of ABA. The early response
genes have been known to encode transcription factors that
activate downstream delayed response genes (Zhu 2002).
Although, specific branches and components exist (Lee
et al. 2001), the signaling pathways for salt, drought, and
Plant Cell Rep (2008) 27:411–424
cold stresses all interact with ABA, and even converge at
multiple steps (Xiong et al. 1999). Abiotic stress signaling
in plants involves receptor-coupled phospho-relay, phosphoionositol-induced Ca2+ changes, mitogen activated
protein kinase (MAPK) cascade, and transcriptional activation of stress responsive genes (Xiong and Zhu 2001).
A number of signaling components are associated with the
plant response to high temperature, freezing, drought and
anaerobic stresses (Grover et al. 2001).
One of the merits for the manipulation of signaling
factors is that they can control a broad range of downstream events that can result in superior tolerance for
multiple aspects (Umezawa et al. 2006). Alteration of
these signal transduction components is an approach to
reduce the sensitivity of cells to stress conditions, or such
that a low level of constitutive expression of stress genes
is induced (Grover et al. 1999). Overexpression of functionally conserved At-DBF2 (homolog of yeast DBf2
kinase) showed striking multiple stress tolerance in
Arabidopsis plants (Lee et al. 1999). Pardo et al. (1998)
also achieved salt stress-tolerant transgenic plants by overexpressing calcineurin (a Ca2+/Calmodulin dependent
protein phosphatase), a protein phosphatase known to be
involved in salt-stress signal transduction in yeast.
Transgenic tobacco plants produced by altering stress
signaling through functional reconstitution of activated
yeast calcineurin not only opened-up new routes for study
of stress signaling, but also for engineering transgenic
crops with enhanced stress tolerance (Grover et al. 1999).
Overexpression of an osmotic-stress-activated protein
kinase, SRK2C resulted in a higher drought tolerance in
A. thaliana, which coincided with the upregulation of
stress-responsive genes (Umezawa et al. 2004). Similarly,
a truncated tobacco mitogen-activated protein kinase
kinase kinase (MAPKKK), NPK1, activated an oxidative
signal cascade resulting in cold, heat, salinity and drought
tolerance in transgenic plants (Kovtun et al. 2000; Shou
et al. 2004). However, suppression of signaling factors
could also effectively enhance tolerance to abiotic stress
(Wang et al. 2005). This hypothesis was based on previous
reports indicating that a and b subunits of farnesyltransferase ERA1 functions as a negative regulator of ABA
signaling (Cutler et al. 1996; Pei et al. 1998). Conditional
antisense downregulation of a or b subunits of protein
farnesyl transferase, resulted in enhanced drought tolerance of Arabidopsis and canola plants.
Choice of promoters
An important aspect of transgenic technology is the regulated expression of transgenes. Tissue specificity of
transgene expression is also an important consideration
417
while deciding on the choice of the promoter so as to
increase the level of expression of the gene. Thus, the
strength of the promoter and the possibility of using stressinducible, developmental-stage-, or tissue-specific promoters have also proved to be critical for tailoring plant
response to these stresses (Bajaj et al. 1999). Some gene
products are needed in large amounts, such as LEA3,
thereby necessitating the need for a very strong promoter.
With other gene products, such as enzymes for polyamine
biosynthesis, it may be better to use an inducible promoter
of moderate strength. The promoters that have been most
commonly used in the production of abiotic stress tolerant
plants so far, include the CaMV 35S, ubiquitin 1 and actin
promoters. These promoters being constitutive in nature,
by and large express the downstream transgenes in all
organs and at all the stages. However, constitutive overproduction of molecules, such as trehalose (Romero et al.
1997) or polyamines (Capell et al. 1998) causes abnormalities in plants grown under normal conditions. Also, the
production of the above-described molecules can be metabolically expensive. In these cases, the use of a stress
inducible promoter may be more desirable. In plants, various types of abiotic stresses induce a large number of wellcharacterized and useful promoters. An ideal inducible
promoter should not only be devoid of any basal level of
gene expression in the absence of inducing agents, but the
expression should be reversible and dose-dependent. The
transcriptional regulatory regions of the drought-induced
and cold-induced genes have been analyzed to identify
several cis-acting and trans-acting elements involved in the
gene expression that is induced by abiotic stress (Shinwari
1999). Most of the stress promoters contain an array of
stress-specific cis-acting elements that are recognized by
the requisite transcription factors; for example, the transcriptional regulation of hsp genes is mediated by the core
‘‘heat shock element’’ (HSE) located in the promoter
region of these genes, 5’ of the TATA box. All the plant
hsp genes sequenced so far have been shown to contain
partly overlapping multiple HSEs proximal to TATA
motif. Apart from these hsp promoters, rd29 and adh gene
promoters induced by osmotic stress and anaerobic stress,
respectively, have also been studied. The Arabidopsis
rd29A and rd29B are stress responsive genes, but are differentially induced under abiotic stress conditions. The
rd29A promoter includes both DRE and ABRE elements,
where dehydration, high salinity and low temperatures
induce the gene, while the rd29B promoter includes only
ABREs and the induction is ABA-dependent. Overexpression of DREB1A transcription factors under the
control of stress inducible promoter from rd29A showed a
better phenotypic growth of the transgenic plants than the
ones obtained using the constitutive CaMV 35S promoter
(Kasuga et al. 1999). A stress inducible expression of
123
418
Arabidopsis CBF1 in transgenic tomato was achieved using
the ABRC1 promoter from barley HAV22 (Lee et al. 2003).
Gene expression is induced by the binding of DREB1A,
which in itself is induced by cold and water stress, to a cisacting DRE element in the promoters of genes such as
rd29A, rd17, cor6.6, cor15A, erd10, and kin1, thereby,
initiating synthesis of gene products imparting tolerance to
low temperatures and water stress in plants. The regions of
respiratory alcohol dehydrogenase adh1 gene promoter in
maize and rice that are required for anaerobic induction
include a string of bases called anoxia response element
(ARE) with the consensus sequence of its core element as
TGGTTT. Besides, other stress-responsive cis-acting promoter sequences like low temperature responsive elements
(LTRD) with a consensus sequence of A/GCCGAC have
been identified in genes such as Cor 6.6, Cor 15 and Cor 78
These basic findings on stress promoters have led to a
major shift in the paradigm for genetically engineering
stress tolerant crops (Katiyar-Aggarwal et al. 1999).
Physiological evaluation of stress effect
A large number of studies have evaluated different transgenic constructs in different plant species, and to different
stresses such as drought, salinity and cold. The expression
of the genes inserted as well as altered levels of metabolites
have been reported in great detail. However, less detail is
given with regard to the methods used to evaluate the stress
response. Although, the transgenic construct is usually
reported to have increased the tolerance to drought in most
of the instances, it is then referred to as such in other
papers. This lack of details applies mostly to drought stress,
the protocols used for salt stress are usually better described (Tarczynsky et al. 1993; Holstrom et al. 2000),
although the levels of salt stress used in some studies are
far beyond what is found in a natural environment. It is
understood that most of these studies are intended to assess
the gene expression, often in model plants, under a particular stress, and extreme situation of stress are often used
to ensure the gene expression. However, these studies may
bring about some misleading conclusions from an agronomic or physiology perspective, where the assessment of
stress tolerance of transgenics needs to be done with
respect to its cross-talk with other stress-related genes/
mechanisms and where the effects of stress need to be
observed over longer periods/conditions. This is particularly important, in order to closely mimic the life span of
most crops under cycles of stress, rather than short exposure to very severe stresses, although we agree that short
exposures to stress are certainly adequate if the purpose is
to assess gene expression only. Therefore, in the following
discussion, we focus on the agronomic/physiological
123
Plant Cell Rep (2008) 27:411–424
perspective and don’t mean to challenge the quality of the
work done to assess gene expression. Our intention is to try
to reconcile both approaches (molecular and agronomic)
toward a common focus: breeding.
Two major issues that typically need to be addressed in
stress response evaluation of transgenics include: (1)
Means of stress imposition, details about the stress, and
growth conditions (including the intensity, timing, and
quickness of imposition, etc.), and (2) ‘‘Hard’’ data on the
response of tested materials to support conclusions (comparison within the same species). Besides, precise details
about the protocols used to evaluate the performance of
plants to any given stress are very essential to assess the
performance of materials.
Means of stress impositions, growth conditions,
and evaluations
Stress conditions used to evaluate the transgenic material in
most of the reports so far, are usually too severe (Nanjo
et al. 1999a; Shinwari et al. 1998; Garg et al. 2002) as
plants are very unlikely to undergo such stresses under field
conditions. Also, the means of evaluation are often significantly different from natural conditions. For example,
Pellegrineschi et al. (2004) compared the performance of
initial events of DREB1A transgenic wheat to the wild
parent by withholding water to 2-week-old seedlings grown
in 5 cm 9 5 cm pots, and then re-watering until maturity
when they were evaluated. Untransformed plants were
nearly dead within 10–15 days of stress imposition, likely
because of a different pattern of water use, whereas
transgenic plants survived in these small pots and ‘‘passed’’
the evaluation successfully; such conditions would obviously not occur in the field. Besides, the type of systems
used to assess plant performance, one would expect the
evaluation to be made, at least, on the basis of biomass
accumulated during the stress.
While the use of PEG (polyethylene glycol) in hydroponics can be useful to test certain response of plants under
a given osmotic potential as reported by Pilon-Smits et al.
(1996, 1999), it offers relatively different conditions than
in the soil where the water reservoir is by definition finite.
Here, the observation on improved growth was explained
by an increased water uptake under the water potential
applied, due to osmolyte production by the transgenic
plant. This is quite possible in such a system because the
water reservoir is unlimited in hydroponics, and because
the water potential is constant. Under soil conditions,
however, the volume of soil surrounding the root where
water can be extracted is limited, and the water potential of
that soil quickly declines upon water uptake by roots,
reaching soil water potential where even the enhanced
Plant Cell Rep (2008) 27:411–424
osmolyte production of the transgenics would be unable to
extract any significant additional amount of water. A more
realistic test of the ability to take up water using osmotic
potential-enhanced transgenics would be to compare their
capacity to extract water from a soil system rather than a
hydroponic system. A recent study by Sivamani et al.
(2000) reported an increased WUE in the transgenic wheat.
Unfortunately there was no control over the soil evaporation that probably accounts for most of the water loss and
explained the very low values of WUE observed. Besides,
investigating drought responses by using fresh weight (Sun
et al. 2001) and other indirect estimates of performance
like growth rate, stem elongation (Pilon-Smits et al. 1995;
Lee et al. 2003), or survival (Pardo et al. 1998) are likely to
give inconsistent results. While applying a drought stress, it
is important to know the stages of drought stress that the
plants are exposed to, for which, a detailed description of
growth conditions, plant size, container size, water availability, and transpiration is needed. It is also crucial to
report the dry weight of tested plants, possibly before and
after the stress period.
Similarly, often the stress imposed has been modified
from 2 days, to 2 weeks, and even 4 weeks using the same
experimental conditions (Lee et al. 2003), without indicating the water holding capacity of the potting mixture
used as well as the plant density. This obviously leads to
different types of stresses, where the plants exposed for
2 days of water stress may well have remained in stage I
when water is abundant (see below), while plants exposed
to 4 weeks stress may have spent most of the time under
stage III where roots may have exhausted all the available
water. Also there are cases where a given quantity of water
is applied to the plants on alternate days from 2 to
10 weeks (Sivamani et al. 2000), thereby, disregarding the
fact that the water requirements increase dramatically
during the period, and probably exposing their plants to an
initial flooding before a severe stress.
419
shoot and stomata progressively close to adjust the water
loss to the water supply, so that leaf turgor is maintained. In
stage III, roots have exhausted all the water available for
transpiration. Stomata are closed and virtually all the
physiological processes contributing to growth, including
photosynthesis are inhibited. This has been used to design
dry-down experiments where the response of plants to
drought is taken as a function of the fraction of soil
moisture available to plant (fraction of transpirable soil
water, FTSW), and not as a function of number of days
after which the stress has been imposed. The former allows
a precise comparison of stress imposed across experiments
and environmental conditions, whereas referring to stress
intensity on the number of days of exposure to stress,
without referring to pot size, evaporative demand, etc., can
lead to erratic and irreproducible data. Based on transpiration values, it is possible to partially compensate the
water loss to apply a milder stress condition, which allows
plants of different sizes to be exposed to a similar drought
stress. For instance, plants exposed to water stress are
allowed to lose a maximum of 70 g per day. Any water loss
in excess of this value is added back on a plant basis. This
allows maintaining the volumetric soil moisture content, a
proxy for water stress, similar in all pots. Amount of daily
water loss can be adapted to increase/decrease the level of
stress. This protocol has the advantage of mimicking the
situation a plant would face in the field, i.e. a progressive
soil drying. This method has been successfully used at
ICRISAT to assess the response of 14 transgenic events of
groundnut (Fig. 1) with rd29A promoter-driven DREB1A
Adequate protocols to apply drought and salinity stress
Unlike what seems to be a common practice in transgenic
evaluation, applying drought does not consist simply in
withholding water. Indeed, we cannot investigate drought
responses of plants without understanding the different
phases that a plant undergoes under drought in natural
conditions. These steps have been described earlier
(Ritchie 1982; Sinclair and Ludlow 1986). In phase I, water
is abundant and plant can take up all the water required by
transpiration and stomata are fully open. During that stage,
the water loss is mostly determined by the environmental
conditions to which the leaves are exposed. During stage II,
the roots are no longer able to supply sufficient water to the
Fig. 1 A typical response curve of groundnut cultivar JL 24 to soildrying condition. This is used to design dry-down experiments where
the response of plants to drought is taken as a function of the fraction
of soil moisture available to plant (fraction of transpirable soil water,
FTSW), and not as a function of the number of days after which the
stress has been imposed
123
420
under contained greenhouse conditions (Bhatnagar-Mathur
et al. 2004, 2006).
Regarding salinity, most of the evaluations reported so
far have been carried out at the seedling stage (Maliro
et al. 2004), although this type of evaluation has been
reported to have little correspondence, if any, with how
plants will later perform under salt stress (Munns et al.
2002; Vadez et al. 2006). Besides, evaluations are made
on a short-term basis by using high concentrations of
salt; way above those found even in highly saline natural
environment that obviously magnifies the effect of
transgenics engineered to excrete salt. Therefore, protocols that use too severe concentrations of salt should be
avoided. A few other subjects of contention include the
treatments that are used as salt stress, and also the
hypothesis about the major determinants of salt stress
tolerance.
It is often assumed that the avoidance of Na+ accumulation and toxicity confers salt tolerance in plants.
Therefore, most of the transgenic work has dealt with genes
involved in Na+ extrusion from the root or Na compartmentation in the vacuoles. However, severe stresses (over
200–300 mM) in hydroponics (Behnam et al. 2006;
Holmstrom et al. 2000; Lee et al. 2003) that are unlikely to
occur in the natural environment will necessarily highlight
those transgenics that are able to excrete Na+ and able to
maintain homeostasis, even though it may be for a short
while. Whether such a strategy is adequate is still an open
question. Vadez et al. (2007) reported that salinity tolerance was not related to differences in the accumulation of
Na+ in chickpea, thereby, a strategy of Na+ excretion in
chickpea would appear inadequate and similar converging
data has been observed in sorghum and millet (unpublished
data).
Procedures for the salinity evaluation of crops are being
optimized to be carried out in soil conditions in an outdoor
facility under natural conditions at ICRISAT. Here, salt
stress is applied to the soil during the early stages of germination and plant development using a staggered salt
application (total amount split in three applications) to
avoid an osmotic shock. Besides, plants are maintained
close to 80% field capacity until maturity to avoid a
possible increase in salt concentration if water is not
replenished regularly. The plant tolerance to stress is
evaluated based on the seed yield since no correlation
between the shoot biomass and seed yield under salinity
has been observed (Vadez et al. 2007). It is likely that
reproduction is the key physiological process affected by
salinity. Therefore, transgenic research intended to
improve salt tolerance should probably be focused on those
processes that appear to be sensitive. A thorough investigation of these processes can only help in devising a
suitable and focused transgenic approach.
123
Plant Cell Rep (2008) 27:411–424
Conclusions
This review summarizes the recent efforts to improve
abiotic stress tolerance in crop plants by employing some
of the stress-related genes and transcription factors that
have been cloned and characterized. The following general
conclusions emerge from this review:
1.
2.
3.
4.
5.
The use of transgenes to improve the tolerance of crops
to abiotic stresses remains an attractive option.
Options targeting multiple gene regulation appear
better than targeting single genes.
An important issue to address is how the tolerance to
specific abiotic stress is assessed, and whether the
achieved tolerance compares to existing tolerance. The
biological cost of production of different metabolites
to cope with stress and their effect on yield should be
properly evaluated.
A well focused approach combining the molecular,
physiological and metabolic aspects of abiotic stress
tolerance is required for bridging the knowledge gaps
between short- and long-term effects of the genes and
their products, and between the molecular or cellular
expression of the genes and the whole plant phenotype
under stress.
Thorough understanding of the underlying physiological processes in response to different abiotic stresses
can efficiently/successfully drive the choice of a
given promoter or transcription factor to be used for
transformation.
Acknowledgments The authors thank Dr. David Hoisington for his
helpful comments on this review. KKS would like to thank the IndoSwiss collaboration in Biotechnology that is jointly funded by the
Department of Biotechnology, Government of India and the Swiss
Development Corporation for providing the funds.
References
Abebe T, Guenzi AC, Martin B, Cushman JC (2003) Tolerance of
Mannitol-accumulating transgenic wheat to water stress and
salinity. Plant Physiol 131:1748–1755
Aharoni A, Dixit S, Jetter R, Thoenes E, van Arkel G, Pereira A
(2004) The SHINE clade of AP2 domain transcription factors
activates wax biosynthesis, alters cuticle properties, and confers
drought tolerance when overexpressed in Arabidopsis. Plant Cell
16:2463–2480
Alia H, Sakamoto A, Murata N (1998) Enhancement of tolerance of
Arabidopsis to high temperature by genetic engineering of the
synthesis of glycine betaine. Plant J 16:155–161
Alia H, Kondo Y, Sakamoto A, Nonaka H, Hayashi H, Pardha Saradhi
P, Chen THH, Norio M (1999) Enhanced tolerance to light stress
of transgenic Arabidopsis plants that express the codA gene for a
bacterial choline oxidase. Plant Mol Biol 40:279–288
Allen RD (1995) Dissection of oxidative stress tolerance using
transgenic plants. Plant Physiol 107:1049–1054
Plant Cell Rep (2008) 27:411–424
Anderson SE, Bastola DR, Minocha SC (1998) Metabolism of
polyamines in transgenic cells of carrot expressing a mouse
ornithine decarboxylase cDNA. Plant Physiol 116:299–307
Apse MP, Aharon GS, Snedden WA, Blumwald E (1999) Salt
tolerance conferred by overexpression of a vascoular Na+/H+
antiport in Arabidopsis. Science 285:1256–1258
Bajaj S, Targolli J, Liu L-F, Ho TH, Wu R (1999) Transgenic
approaches to increase dehydration stress tolerance in plants.
Mol Breed 5:493–503
Bartels D, Sunkar R (2005) Drought and salt tolerance in plants. Crit
Rev Plant Sci 21:1–36
Behnam B, Kikuchi A, Celebi-Toprak F, Yamanaka S, Kasuga M,
Yamaguchi-Shinozaki K, Watanabe KN (2006) The Arabidopsis
DREB1A gene driven by the stress-inducible rd29A promoter
increases salt-stress tolerance in proportion to its copy number in
tetrasomic tetraploid potato (Solanum tuberosum). Plant Biotech
23:169–177
Bhatnagar-Mathur P, Devi M J, Serraj R, Yamaguchi-Shinozaki K,
Vadez V, Sharma KK (2004) Evaluation of transgenic groundnut
lines under water limited conditions. Int Arch Newsl 24:33–34
Bhatnagar-Mathur P, Devi M J, Reddy DS, Vadez V, YamaguchiShinozaki K, Sharma KK (2006) Overexpression of Arabidopsis
thaliana DREB1A in transgenic peanut (Arachis hypogaea L.)
for improving tolerance to drought stress (poster presentation).
In: Arthur M. Sackler Colloquia on ‘‘From Functional Genomics
of Model Organisms to Crop Plants for Global Health’’, April 3–
5, 2006. National Academy of Sciences, Washington, DC
Bohnert HJ, Shen B (1999) Transformation and compatible solutes.
Sci Hort 78:237–260
Bohnert HJ, Nelson DF, Jenson RG (1995) Adaptation to environmental stresses. Plant Cell 7:1099–1111
Bordás M, Montesinos C, Dabauza M, Salvador A, Roig LA, Serrano
R, Moreno V (1997) Transfer of the yeast salt tolerance gene
HAL1 to Cucumis melo L. cultivars and in vitro evaluation of
salt tolerance. Transgenic Res 5:1–10
Bowler C, Slooten L, Vandenbranden S, Rycke RD, Botterman J,
Sybesma C, van Montagu M, Inze D (1991) Manganese
superoxide dismutase can reduce cellular damage mediated by
oxygen radicals in transgenic plants. EMBO J 10:1723–1732
Bray EA (1993) Molecular responses to water deficit. Plant Physiol
103:1035–1040
Burke EJ, Brown SJ, Christidis N (2006) Modeling the recent
evolution of global drought and projections for the twenty-first
century with the Hadley centre climate model. J Hydrometeor
7:1113–1125
Capell T, Escobar C, Lui H, Burtin H, Lepri O, Christou P (1998)
Overexpression of the oat arginine decarboxylase cDNA in
transgenic rice (Oryza sativa L.) affects normal development
patterns in vitro and results in putrescine accumulation in
transgenic plants. Theor Appl Genet 97:246–254
Capell T, Bassie L, Christou P (2004) Modulation of the polyamine
biosynthetic pathway in transgenic rice confers tolerance to
drought stress. Proc Natl Acad Sci USA 101:9909–9914
Chen TH, Murata N (2002) Enhancement of tolerance of abiotic stress
by metabolic engineering of betaines and other compatible
solutes. Curr Opin Plant Biol 5:250–257
Cheng WH, Endo A, Zhou L, Penney J, Chen HC, Arroyo A, Leon P,
Nambara E, Asami T, Seo M (2002) A unique short-chain
dehydrogenase/reductase in Arabidopsis glucose signaling and
abscisic acid biosynthesis and functions. Plant Cell 14:2723–
2743
Chinnusamy V, Jagendorf A, Zhu JK (2005) Understanding and
improving salt tolerance in plants. Crop Sci 45:437–448
Cortina C, Culiá n ez-Maciá F (2005) Tomato abiotic stress enhanced
tolerance by trehalose biosynthesis. Plant Sci 169:75–82
421
Cutler S, Ghassemian M, Bonetta D, Cooney S, McCourt P (1996) A
protein farnesyl transferase involved in abscisic acid signal
transduction in Arabidopsis. Science 273:1239–1241
Delauney AJ, Verma DPS (1993) Proline biosynthesis and osmoregulation in plants. Plant J 4:215–223
Diamant S, Eliahu N, Rosenthal D, Goloubinoff P (2001) Chemical
chaperones regulate molecular chaperones in vitro and in cells
under combined salt and heat stresses. J Biol Chem 276:39586–
39591
Dure L III (1993) A repeating 11-mer amino acid motif and plant
desiccation. Plant J 3:363–369
Dure L III, Crouch M, Harada J, Ho T-HD, Mundy J (1989) Common
amino acid sequence domains among the LEA proteins of higher
plants. Plant Mol Biol 12:475–486
FAO (Food, Agriculture Organization of the United Nations) (2004)
FAO production yearbook. FAO, Rome
Fukai S, Cooper M (1995) Developing resistant cultivars using
physiomorphological traits in rice. Field Crops Res 40:67–86
Galau GA, Bijaisoradat N, Hughes DW (1987) Accumulation kinetics
of cotton late embryogenesis-abundent (Lea) mRNAs and
storage protein mRNAs: coordinate regulation during embryogenesis and role of abscisic acid. Dev Biol 123:198–212
Gao M, Sakamoto A, Miura K, Murata N, Sugiura A, Tao R (2000)
Transformation of Japanese persimmon (Diospyros kaki Thunb.)
with a bacterial gene for choline oxidase. Mol Breed 6:501–510
Garg AK, Kim JK, Owens TG, Ranwala AP, Choi YC, Kochian LV,
Wu RJ (2002) Trehalose accumulation in rice plants confers high
tolerance levels to different abiotic stresses. Proc Natl Acad Sci
USA 99:15898–15903
Gisbert C, Rus AM, Bolarin MC, Lopez-Coronado M, Arrillaga I,
Montesinos C, Caro M, Serrano R, Moreno V (2000) The yeast
HAL1 gene improves salt tolerance of transgenic tomato. Plant
Physiol 123:393–402
Goyal K, Walton LJ, Tunnacliffe A (2005) LEA proteins prevent
protein aggregation due to water stress. Biochem J 388:151–157
Grover A, Minhas D (2000) Towards production of abiotic stress
tolerant transgenic rice plants: issues, progress and future
research needs. Proc Indian Natl Sci Acad B Rev Tracts. Biol
Sci 66:13–32
Grover A, Sahi C, Sanan N, Grover A (1999) Taming abiotic stresses
in plants through genetic engineering: current strategies and
perspective. Plant Sci 143:101–111
Grover A, Kapoor A, Satya Lakshmi O, Agrawal S, Sahi C, KatiyarAgarwal S, Agarwal M, Dubey H (2001) Understanding
molecular alphabets of the plant abiotic stress responses. Curr
Sci 80:206–216
Guiltinan MJ, Marcotte WR, Quatrano RS (1990) A plant leucine
zipper protein that recognizes an abscisic acid response element.
Science 250:267–271
Haake V, Cook D, Riechmann JL, Pineda O, Thomashow MF, Zhang
JZ (2002) Transcription factor CBF4 is a regulator of drought
adaptation in Arabidopsis. Plant Physiol 130:639–648
Hare PD, Cress WA, Van Staden J (1998) Dissecting the roles of
osmolyte accumulation during stress. Plant Cell Environ 21:535–
553
Hayashi H, Mustardy L, Deshnium P, Ida M, Murata N (1997)
Transformation of Arabidopsis thaliana with the codA gene for
choline oxidase: accumulation of glycine betaine and enhanced
tolerance to salt and cold stress. Plant J 12:133–142
Hayashi H, Alia, Sakamoto A, Nonaka H, Chen THH, Murata N
(1998) Enhanced germination under high-salt conditions of seeds
of transgenic Arabidopsis with a bacterial gene (codA) for
choline oxidase. J Plant Res 111:357–362
Holmstrom KO, Manty E, Welin B, Palva ET (1996) Drought
tolerance in tobacco. Nature 379:683–684
123
422
Holmstrom KO, Somersalo S, Mandal A, Palva ET, Welin B (2000)
Improved tolerance to salinity and low temperature in transgenic
tobacco producing glycine betaine. J Expt Bot 51:177–185
Hsieh TH, Lee JT, Charng YY, Chan MT (2002) Tomato plants
ectopically expressing Arabidopsis CBF1 show enhanced resistance to water deficit stress. Plant Physiol 130:618–626
Ishitani M, Xiong L, Stevenson B, Zhu J-K (1997) Genetic analysis of
osmotic and cold stress signal transduction in Arabidopsis:
interactions and convergence of abscisic acid-dependent and
abscisic acid-independent pathways. Plant Cell 9:1935–1949
Iwasaki T, Kiyosue T, Yamaguchi-Shinozaki K (1997) The dehydration-inducible rd17 (cor47) gene and its promoter region in
Arabidopsis thaliana. Plant Physiol 115:128
Jaglo KR, Kleff KL, Amundsen X, Zhang V, Haake JZ, Zhang T,
Thomashow MF (2001) Components of the Arabidopsis Crepeat/dehydration-responsive element binding factor coldresponse pathway are conserved in Brassica napus and other
plant species. Plant Physiol 127:910–917
Jaglo-Ottosen KR, Gilmour SJ, Zarka DG, Schabenberger O,
Thomashow MF (1998) Arabidopsis CBF1 overexpression
induces cor genes and enhances freezing tolerance. Science
280:104–106
Kagaya Y, Hobo T, Murata M, Ban A, Hattori T (2002) Abscisic
acidinduced transcription is mediated by phosphorylation of an
abscisic acid response element binding factor, TRAB1. Plant
Cell 14:3177–3189
Kasuga M, Liu Q, Miura S, Yamaguchi-Shinozaki K, Shinozaki K
(1999) Improving plant drought, salt, and freezing tolerance by
gene transfer of a single stress inducible transcription factor. Nat
Biotechnol 17:287–291
Kasuga M, Miura S, Shinozaki K, Yamaguchi-Shinozaki K (2004) A
combination of the Arabidopsis DREB1A gene and stressinducible rd29A promoter improved drought- and low-temperature stress tolerance in tobacco by gene transfer. Plant Cell
Physiol 45:346–350
Katiyar-Agarwal S, Agarwal M, Grover A (1999) Emerging trends in
agricultural biotechnology research: use of abiotic stress induced
promoter to drive expression of a stress resistance gene in the
transgenic system leads to high level stress tolerance associated
with minimal negative effects on growth. Curr Sci 77:1577–1579
Katiyar-Agarwal S, Agarwal M, Grover A (2003) Heat-tolerant
basmati rice engineered by over-expression of hsp101. Plant Mol
Biol 51:677–686
Kimpel JA, Key JL (1985) Heat shock in plants. Trends Biochem Sci
10:353–357
Kovtun Y, Chiu WL, Tena G, Sheen J (2000) Functional analysis of
oxidative stress-activated mitogen-activated protein kinase cascade in plants. Proc Natl Acad Sci USA 97:2940–2945
Kumria R, Rajam MV (2002) Ornithine decarboxylase transgene in
tobacco affects polyamines, in vitro-morphogenesis and response
to salt stress. J Plant Physiol 159:983–990
Lee JH, Hubel A, Schoffl F (1995) Derepression of activity of
genetically engineered heat-shock factor causes constitutive
synthesis of heat shock proteins and increased thermotolerance
in transgenic Arabidopsis. Plant J 8:603–612
Lee JH, van Montagu M, Verbruggen N (1999) A highly conserved
kinase is an essential component for stress tolerance in yeast and
plant cells. Proc Natl Acad Sci USA 96:5873–5877
Lee H, Xiong L, Gong Z, Ishitani M, Stevenson B, Zhu JK (2001) The
Arabidopsis HOS1 gene negatively regulates cold signal transduction and encodes a RING-finger protein that displays coldregulated nucleo-cytoplasmic partitioning. Gene Dev 15:912–924
Lee SS, Cho HS, Yoon GM, Ahn JW, Kim HH, Pai HS (2003)
Interaction of NtCDPK1 calcium-dependent protein kinase with
NtRpn3 regulatory subunit of the 26S proteasome in Nicotiana
tabacum. Plant J 33:825–840
123
Plant Cell Rep (2008) 27:411–424
Li HY, Chang CS, Lu LS, Liu CA, Chan MT, Chang YY (2003)
Over-expression of Arabidopsis thaliana heat shock factor gene
(AtHsfA1b) enhances chilling tolerance in transgenic tomato. Bot
Bull Acad Sin 44:129–140
Lilius G, Holmberg N, Bülow L (1996) Enhanced NaCl stress
tolerance in transgenic tobacco expressing bacterial choline
dehydrogenase. Biotechnol 14:177–180
Lindquist S (1986) The heat-shock response. Annu Rev Biochem
55:1151–1191
Liu Q, Kasuga M, Sakuma Y, Abe H, Miura S, Yamaguchi-Shinozaki
K, Shinozaki K (1998) Two transcription factors, DREB1 and
DREB2, with an EREBP/AP2 DNA binding domain separate
two cellular signal transduction pathways in drought- and low
temperature-responsive gene expression, respectively, in
Arabidopsis. Plant Cell 10:1391–1406
Malik MK, Solvin JP, Hwang CH, Zimmerman JL (1999) Modified
expression of a carrot small heat-shock protein gene, Hsp 17.7
results in increased or decreased thermotolerance. Plant J 20:89–
99
Maliro MFA, McNeil D, Kollmorgen J, Pittock C, Redden B (2004)
Screening chickpea (Cicer arietinum L.) and wild relatives
germplasm from diverse sources for salt tolerance. New
directions for a diverse planet: Proceedings of the 4th International Crop Science Congress, Brisbane, Australia, 26 Sep to 1
Oct 2004
Mckersie BD, Bowley SR, Harjanto E, Leprince O (1996) Water deficit
tolerance and field performance of transgenic alfalfa overexpressing superoxide dismutase. Plant Physiol 111:1177–1181
McNeil SD, Nuccio ML, Hanson AD (1999) Betaines and related
osmoprotectants. Targets for metabolic engineering of stress
resistance. Plant Physiol 120:945–949
McNeil SD, Nuccio ML, Rhodes D, Shachar-Hill Y, Hanson AD
(2000) Radiotracer and computer modeling evidence that
phosphobase methylation is the main route of choline synthesis
in tobacco. Plant Physiol 123:371–380
Miroshnichenko S, Tripp J, Nieden UZ, Neumann D, Conrad U,
Manteuffel R (2005) Immunomodulation of function of small
heat shock proteins prevents their assembly into heat stress
granules and results in cell death at sublethal temperatures. Plant
J 41:269–281
Mundy J, Chua N-H (1988) Abscisic acid and water-stress induce the
expression of a novel rice gene. EMBO J 7:2279–2286
Mundy J, Yamaguchi-Shinozaki K, Chua NH (1990) Nuclear proteins
bind conserved elements in the abscisic acid-responsive promoter of a rice rab gene. Proc Natl Acad Sci USA 87:406–410
Munns R, Husain S, Rivelli AR, James RA, Condon AG, Lindsay
MP, Lagudah ES, Schachtman DP, Hare RA (2002) Avenues for
increasing salt tolerance of crops, and the role of physiologically
based selection traits. Plant Soil 247:93–105
Murakami Y, Tsuyama M, Kobayashi Y, Kodama H, Iba K (2000)
Trienoic fatty acids and plant tolerance of high temperature.
Science 287:476–479
Murata N, Ishizaki-Nishizawa O, Higashi S, Hayashi S, Tasaka Y,
Nishida I (1992) Genetically engineered alteration in the chilling
sensitivity of plants. Nature 356:710–713
Nanjo T, Kobayashi M, Yoshiba Y, Kakubari Y, YamaguchiShinozaki K, Shinozaki K (1999a) Antisense supression of
proline degradation improves tolerance to freezing and salinity
in Arabidopsis thaliana. FEBS Lett 461:205–210
Nordin K, Heino P, Palva ET (1991) Separate signal pathways
regulate the expression of a low-temperature-induced gene in
Arabidopsis thaliana (L.) Heynh. Plant Mol Biol 115:875–879
Oberschall A, Deak M, Torok K, Sass L, Vass I, Kovacs I, Feher A,
Dudits D, Hovarth GV (2000) A novel aldose/aldehyde reductase
protects transgenic plants against lipid peroxidation under
chemical and drought stress. Plant J 24:437–446
Plant Cell Rep (2008) 27:411–424
Ozturk ZN, Talame V, Deyholos M, Michalowski CB, Galbraith DW,
Gozukirmizi N, Tuberosa R, Bohnert HJ (2002) Monitoring
large-scale changes in transcript abundance in drought- and saltstressed barley. Plant Mol Biol 48:551–573
Passioura J (1977) Physiology of grain yield in wheat growing on
stored water. Aust J Plant Physiol 3:559–565
Passioura J (2007) The drought environment: physical, biological and
agricultural perspectives. J Exp Bot 58:113–117
Pardo JM, Reddy MP, Yang S (1998) Stress signaling through Ca2+/
Calmodulin dependent protein phosphatase calcineurin mediates
salt adaptation in plants. Proc Natl Acad Sci USA 95:9681–9683
Pei ZM, Ghassemian M, Kwak CM, McCourt P, Schroeder JI (1998)
Role of farnesyltransferase in ABA regulation of guard cell
anion channels and plant water loss. Science 282:287–290
Pellegrineschi A, Reynolds M, Pacheco M, Brito R M, Almeraya R,
Yamaguchi-Shinozaki K, Hoisington D (2004) Stress-induced
expression in wheat of the Arabidopsis thaliana DREB1A gene
delays water stress symptoms under greenhouse conditions.
Genome 47:493–500
Perl A, Perl-Treves R, Galili S, Aviv D, Shalgi E, Malkin S, Galun E
(1993) Enhanced oxidative stress defense in transgenic potato
expressing tomato Cu, Zn superoxide dismutases. Theor Appl
Genet 85:568–576
Pilon-Smits EAH, Ebskamp MJM, Paul MJ, Jeuken JW, Weisbeek,
Smeekens SCM (1995) Improved performance of transgenic
fructan-accumulating tobacco under drought stress. Plant Physiol
107:125–130
Pilon-Smits EAH, Ebskamp MJM, Jeuken MJW, van der Meer IM,
Visser RGF, Weisbeek PJ, Smeekens JCM (1996) Microbial
fructan production in transgenic potato plants and tubers. Ind
Crops Prod 5:35–46
Pilon-Smits EAH, Terry N, Sears T, Kim H, Zayed A, Hwang S, Van
Dun K, Voogd E, Verwoerd TC, Krutwagen RWHH, Giddijn JM
(1998) Trehalose-producing transgenic tobacco plants show
improved growth performance under drought stress. J Plant
Physiol 152:525–532
Pilon-Smits EAH, Terry N, Sears T, van Dun K (1999) Enhanced
drought resistance in fructan-producing sugar beet. Plant Physiol
Biochem 37:313–317
Quimlo CA, Torrizo LB, Setter TL, Ellis M, Grover A, Abrigo EM,
Oliva NP, Ella ES, Carpena AL, Ito O, Peacock WJ, Dennis E,
Datta SK (2000) Enhancement of submergence tolerance in
transgenic rice plants overproducing pyruvate decarboxylase.
J Plant Physiol 156:516–521
Quintero FJ, Blatt MR, Pardo JM (2000) Functional conservation
between yeast and plant endosomal Na(+)/H(+) antiporters.
FEBS Lett 471:224–228
Ritchie GA (1982) Carbohydrate reserves and root growth potential in
Douglas-fir seedlings before and after cold storage. Can J For
Res 12:905–912
Rohila JS, Jain RK, Wu R (2002) Genetic improvement of Basmati
rice for salt and drought tolerance by regulated expression of a
barley Hva1 cDNA. Plant Sci 163:525–532
Romero C, Belles JM, Vaya JL, Serrano R, Culianez-Macia FA
(1997) Expression of the yeast trehalose-6-phosphate synthase
gene in transgenic tobacco plants: pleiotropic phenotypes
include drought tolerance. Planta 201:293–297
Roxas VP, Smith RK Jr, Allen ER, Allen RD (1997) Overexpression
of glutathione S-transferase/glutathione peroxidase enhances the
growth of transgenic tobacco seedlings during stress. Nat
Biotechnol 15:988–991
Roy M, Wu R (2001) Arginine decarboxylase transgene expression
and analysis of environmental stress tolerance in transgenic rice.
Plant Sci 160:869–875
Roy M, Wu R (2002) Overexpression of S-adenosylmethionine
decarboxylase gene in rice increases polyamine level and
423
enhances sodium chloride-stress tolerance. Plant Sci 163:987–
992
Rubio MC, González EM, Minchin FR, Webb KJ, Arrese-Igor C,
Ramos J, Becana M (2002) Effects of water stress on antioxidant
enzymes of leaves and nodules of transgenic alfalfa overexpressing superoxide dismutases. Physiol Plant 115:531–540
Sakamoto A, Alia, Murata N (1998) Metabolic engineering of rice
leading to biosynthesis of glycine betaine and tolerance to salt
and cold. Plant Mol Biol 38:1011–1019
Sakamoto A, Valverde R, Alia, Chen TH, Murata N (2000)
Tranformation of Arabidopsis with the codA gene for choline
oxidase enhances freezing tolerance of plants. Plant J 22:449–
453
Sakuma Y, Maruyama K, Osakabe Y, Qin F, Seki M, Shinozaki K,
Yamaguchi-Shinozaki K (2006) Functional analysis of an
Arabidopsis transcription factor, DREB2A, involved in droughtresponsive gene expression. The Plant Cell 18:1292–1309
Schobert B, Tschesche H (1978) Unusual solution properties of
proline and its interaction with proteins. Biochem Biophys Acta
541:270–277
Seki M, Narusaka M, Abe H, Kasuga M, Yamaguchi- Shinozaki K,
Carninci P, Hayashizaki Y, Shinozaki K (2001) Monitoring the
expression pattern of 1300 Arabidopsis genes under drought and
cold stresses by using a full-length cDNA Microarray. Plant Cell
13:61–72
Sen Gupta A, Heinen JL, Holady AS, Burke JJ, Allen RD (1993)
Increased resistance to oxidative stress in transgenic plants that
over-express chloroplastic Cu/Zn superoxide dismutase. Proc
Nat Acad Sci USA 90:1629–1633
Serraj R, Sinclair TR (2002) Osmolyte accumulation: can it really
help increase crop under drought conditions? Plant Cell Environ
25:333–341
Shen B, Jensen RG, Bohnert HJ (1997) Increased resistance to
oxidative stress in transgenic plants by targeting mannitol
biosynthesis to chloroplasts. Plant Physiol 113:1177–1183
Shi H, Ishitani M, Kim C, Zhu JK (2000) The Arabidopsis thaliana
salt tolerance gene SOS1 encodes a putative Na+/H+ antiporter.
Proc Nat Acad Sci USA 97:6896–6901
Shinozaki K, Yamaguchi-Shinozaki K (1997) Gene expression and
signal transduction in water-stress response. Plant Physiol
115:327–334
Shinozaki K, Yamaguchi-Shinozaki K (1999) Molecular responses to
drought stress. In: Shinozaki K, Yamaguchi-Shinozaki K (eds)
Molecular responses to cold, drought, heat and salt stress in
higher plants. R.G. Landes Co., Austin, pp 11–28
Shinwari ZK (1999) Function and regulation of genes that are induced
by dehydration stress. Biosci Agric 5:39–47
Shinwari ZK, Nakashima K, Miura S, Kasuga M, Seki M, Yamaguchi-Shinozaki K, Shinozaki K (1998) An Arabidopsis gene
family encoding DRE/CRT binding proteins involved in lowtemperature-responsive gene expression. Biochem Biophys Res
Commun 50:161–170
Shou H, Bordallo P, Wang K (2004) Expression of the Nicotiana
protein kinase (NPK1) enhanced drought tolerance in transgenic
maize. J Exp Bot 55:1013–1019
Simpson SD, Nakashima K, Narusaka Y, Seki M, Shinozaki K,
Yamaguhci-Shinozaki K (2003) Two different novel cis-acting
elements of erd1, a clpA homologous Arabidopsis gene function
in induction by dehydration stress and dark-induced senescence.
Plant J 33:259–270
Sinclair TR, Ludlow MM (1986) Influence of soil water supply on the
plant water balance of four tropical grain legumes. Aust J Plant
Physiol 13:329–341
Sinclair BJ, Jaco Klok C, Chown SL (2004) Metabolism of the subAntarctic caterpillar Pringleophaga marioni during cooling,
freezing and thawing. J Exp Biol 207:1287–1294
123
424
Sivamani E, Bahieldin A, Wraith JM, Al-Niemi T, Dyer WE, Ho
THD, Qu R (2000) Improved biomass productivity and water use
efficiency under water deficit conditions in transgenic wheat
constitutively expressing the barley HVA1 gene. Plant Sci
155:1–9
Skriver K, Mundy J (1990) Gene expression in response to abscisic
acid and osmotic stress. Plant Cell 5:503–512
Slooten L, Capiau K, Van Camp W, Montagu MV, Sybesma C, Inzé
D (1995) Factors affecting the enhancement of oxidative stress
tolerance in transgenic tobacco overexpressing manganese
superoxide dismutase in the chloroplasts. Plant Physiol
107:737–775
Sun W, Bernard C, van de Cotte B, Montagu MV, Verbruggen N
(2001) At-HSP17.6A, encoding a small heat-shock protein in
Arabidopsis, can enhance osmotolerance upon overexpression.
Plant J 27:407–415
Tarczynski MC, Jensen RG, Bohnert HJ (1993) Stress protection of
transgenic tobacco by production of osmolyte mannitol. Science
259:508–510
Thomashow M F (1998) Role of cold responsive genes in plant
freezing tolerance. Plant Physiol 118:1–7
Turner NC, Shahal A, Berger JD, Chaturvedi SK, French RJ, Ludwig
C, Mannur DM, SJ Singh SJ, Yadava HS (2007) Osmotic
adjustment in chickpea (Cicer arietinum L.) results in no yield
benefit under terminal drought. J Exp Bot 58:187–194
Umezawa T, Yoshida R, Maruyama K, Yamaguchi-Shinozaki K,
Shinozaki K (2004) SRK2C, a SNF1-related protein kinase 2,
improves drought tolerance by controlling stress-responsive gene
expression in Arabidopsis thaliana. Proc Natl Acad Sci
USA.101:17306–17311
Umezawa T, Fujita M, Fujita Y, Yamaguchi-Shinozaki K, Shinozaki
K (2006) Engineering drought tolerance in plants: discovering
and tailoring genes to unlock the future. Curr Opin Biotechnol
17:113–122
Vadez V, Krishnamurthy L, Gaur PM, Upadhyaya HD, Hoisington
DA, Varshney RK, Turner NC, Siddique KHM (2006) Tapping
the large genetic variability for salinity tolerance in chickpea.
Proceeding of the Australian Society of Agronomy meeting
(10–14 Sept 2006) http://www.agronomy.org.au
Vadez V, Krishnamurthy L, Serraj R, Gaur PM, Upadhyaya HD,
Hoisington DA, Varshney RK, Turner NC, Siddique KHM
(2007) Large variation in salinity tolerance in chickpea is
explained by differences in sensitivity at the reproductive stage.
Field Crop Res (in press)
Van Camp W, Capiau K, Van Montagu M, Inzé D, Slooten L (1996)
Enhancement of oxidative stress tolerance in transgenic tobacco
plants overproducing Fe-superoxide dismutase in chloroplasts.
Plant Physiol 112:1703–1714
Vierling E (1991) The roles of heat shock proteins in plants. Annu
Rev Plant Physiol Plant Mol Biol 42:579–620
Vinocur B, Altman A (2005) Recent advances in engineering plant
tolerance to abiotic stress: achievements and limitations. Curr
Opin Biotechnol 16:123–132
Voetberg GS, Sharp RE (1991) Growth of the maize primary root tip
at low water potentials. III. Role of increased proline deposition
in osmotic adjustment. Plant Physiol 96:1125–1130
Vranova E, Inze D, Van Breusegem F (2002) Signal transduction
during oxidative stress. J Exp Bot 53:1227–1236
Waie B, Rajam MV (2003) Effect of increased polyamine
biosynthesis on stress responses in transgenic tobacco by
introduction of human S-adenosylmethionine gene. Plant Sci
164:727–734
Wang Y, Ying J, Kuzma M, Chalifoux M, Sample A, McArthur C,
Uchacz T, Sarvas C, Wan J, Dennis DT et al (2005) Molecular
123
Plant Cell Rep (2008) 27:411–424
tailoring of farnesylation for plant drought tolerance and yield
protection. Plant J 43:413–424
Waters ER, Lee GJ, Vierling E (1996) Evolution, structure and
function of the small heat shock proteins in plants. J Exp Bot
47:325–338
Winicov I, Bastola DR (1997) Salt tolerance in crop plants: new
approaches through tissue culture and gene regulation. Acta
Physiol Plant 19:435–449
Winicov I, Bastola DR (1999) Transgenic overexpression of the
transcription factor Alfin1 enhances expression of the endogenous MsPRP2 gene in alfalfa and improves salinity tolerance of
the plants. Plant Physiol 120:473–480
Xiong L, Zhu JK (2001) Plant abiotic stress signal transduction:
molecular and genetic perspectives. Physiol Plant 112:152–166
Xiong L, Ishitani M, Zhu J-K (1999) Interaction of osmotic stress,
temperature, and abscisic acid in the regulation of gene
expression in Arabidopsis. Plant Physiol 119:205–211
Xu D, Duan X, Wang B, Hong B, Ho T-HD, Wu R (1996) Expression
of a late embryogenesis abundant protein gene, HVA1, from
barley confers tolerance to water deficit and salt stress in
transgenic rice. Plant Physiol 110:249–257
Xu K, Xu X, Fukao T, Canlas P, Maghirang-Rodriguez R (2006)
Sub1A is an ethylene-response-factor–like gene that confers
submergence tolerance to rice. Nature 442:705–708
Yamada M, Morishita H, Urano K, Shiozaki N, Yamaguchi-Shinozaki K, Shinozaki K, Yoshiba Y (2005) Effects of free proline
accumulation in petunias under drought stress. J Exp Bot
56:1975–1981
Yamaguchi-Shinozaki K, Shinozaki K (1993) Characterisation of the
expression of a dessication-responsive rd29 gene of Arabidopsis
thaliana and analysis of its promoter in transgenic plants. Mol
Gen Genet 236:331–340
Yamaguchi-Shinozaki K, Urao T, Iwasaki T, Kiyosue T, Shinozaki K
(1994) Function and regulation of genes that are induced by
dehydration stress in Arabidopsis thaliana. JIRCAS J 1:69–79
Yang G, Rhodes D, Joly RJ (1996) Effects of high temperature on
membrane stability and chlorophyll fluorescence in glycine
betaine-deficient and glycine betaine containing maize lines.
Aust J Plant Physiol 23:437–443
Zhang H-X, Blumwald E (2001) Transgenic salt-tolerant tomato
plants accumulate salt in foliage but not in fruit. Nat Biotechnol
19:765–768
Zhang JZ, Creelman RA, Zhu JK (2004) From laboratory to field.
Using information from Arabidopsis to engineer salt, cold, and
drought tolerance in crops. Plant Physiol 135:615–621
Zhang JY, Broeckling CD, Blancaflor EB, Sledge MK, Sumner LW,
Wang ZY (2005) Overexpression of WXP1, a putative Medicago
truncatula AP2 domain-containing transcription factor gene,
increases cuticular wax accumulation and enhances drought
tolerance in transgenic alfalfa (Medicago sativa). Plant J 42:689–
707
Zhao HW, Chen YJ, Hu YL, Gao Y, Lin ZP (2000) Construction of a
trehalose-6-phosphate synthase gene driven by drought responsive promoter and expression of drought-resistance in transgenic
tobacco. Acta Bot Sinica 42:616–619
Zhu JK (2002) Salt and drought stress signal transduction in plants.
Annu Rev Plant Physiol Plant Mol Biol 53:247–273
Zhu B, Su J, Chang M, Verma DPS, Fan YL, Wu R (1998)
Overexpression of delta1-pyrroline-5-carboxylate synthase gene
and analysis of tolerance to water and salt stress in transgenic
rice. Plant Sci 199:41–48
Zhu L, Tang GS, Hazen SP, Kim HS, Ward RW (1999) RFLP-based
genetic diversity and its development in Shaanxi wheat lines.
Acta Bot Boreali Occident Sin 19:13